1,1'-(1,3-phenylene)bis-1H-pyrrole-2,5-dione

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

Reproductive toxicity (Screeening for reproductive/developmental toxicity): The NOEL and NOAEL for reproductive/developmental toxicity (parental and F1) were considered to be 240 mg/kg/day (OECD 422/GLP).

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 15-10-2010 to 31-03-2011

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study without detailed documentation

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

GLP compliance:

yes

Limit test:

no

Specific details on test material used for the study:

Purity: 99.3%

Species:

rat

Strain:

other: Crl:CD (SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Co., Ltd
- Age at study initiation: 9 weeks
- Weight at study initiation: 476.7± 33.3g (males) and 286.0± 20.2 (female) at Day 0 of test item adminstration
- Fasting period before study: 16 hrs
- Housing: Individual in aluminum front and floor stainless steel mesh breeding cage
- Diet: Radiation sterilized solid feed (CRF-1, Lot No. 100203, 100302, Oriental Yeast Co.) ad libitum
- Water: ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22.8 to 23.2 ° C
- Humidity (%): 53.8 to 64.8%
- Air changes (per hr): 12 times per hr
- Photoperiod (hrs dark / hrs light): 12 hrs light/dark

Route of administration:

oral: gavage

Vehicle:

other: 0.5% methyl cellulose

Details on exposure:

VEHICLE
- Amount of vehicle (if gavage): 10 mL/kg

Details on mating procedure:

- M/F ratio per cage: 1:1
- Length of cohabitation: 14 days
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as day 0 of pregnancy

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

Males: 42 days
Females; 42-52 days (from 14 days before mating to day 4 of lactation)

Frequency of treatment:

Daily

Dose / conc.:

15 mg/kg bw/day (nominal)

Dose / conc.:

60 mg/kg bw/day (nominal)

Dose / conc.:

240 mg/kg bw/day (nominal)

No. of animals per sex per dose:

Males: 7 rats/group (control and high dose groups of main study)
+5 rats/group (control and high dose groups of recovery).
12 rats/group (low and middle dose groups of main study).

Females: 12 rats/group (all groups of main study)
+ 5 rats/group (control and high dose groups of recovery).

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
In the 14-day range finding study with 6 males and 6 females at doses of 0, 20, 60, 200, 600 mg/kg bw/day, following findings were reported.

600 mg/kg bw/day: Death (M6/6, F6/6), diarrhea, mucous stool, soiled fur, ptosis (MF), body weight↓(MF), food consumption↓(MF), enlarged lumen of the stomach, red or black
patches in the stomach, enlarged lumen of the small intestine, liquid content in the stomach and the large intestine, reddish content in the small and large
intestine, atrophy of the thymus, blackish discoloration of the large intestine, brownish discoloration of the lymph node, red patch / edema of the thymus,
enlarged adrenal, emaciation (MF)

200 mg/kg bw/day: Diarrhea, suppressed body weight gain, enlarged lumen of the stomach / small intestine / large intestine, atrophy of the spleen and the thymus (F1/6), Hct↓,
Hgb↓, RBC↓(M), TP↓, Alb↓, BUN↓ (M), monocyte↑(F), red or brown patches in the mesenteric lymph node (MF)

60 mg/kg bw/day: Hct↓, Hgb↓, RBC↓ (M), TP↓, Alb↓, BUN↓(M)

20 mg/kg bw/day: No effect

Parental animals: Observations and examinations:

DETAILED CLINICAL OBSERVATIONS: Yes

BODY WEIGHT: Yes

HAEMATOLOGY: Yes

CLINICAL CHEMISTRY: Yes

URINALYSIS: Yes (males only)

Behaviour: Yes (Functional Observational Battery:detailed observation, sensory reactivities, grasping power, motor activity).

Reproductive indices:

Copulaiton, fertiltiy, gestation, implantation, delivery index.

Offspring viability indices:

Live birth, Viability index on day 4

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Death occurred in 4 females and 2 males at 240 mg/kg bw/day. These animals showed loose stools, diarrhea, abnormal respiration noise, irregular respiration, etc. before death.

In animals that survived, salivation was observed in males in the 60 mg/kg group and in animals of both sexes in the 240 mg/kg group.

Mortality:

mortality observed, treatment-related

Description (incidence):

Death occurred in 2 males and 2 females in the 240 mg/kg group of the main study group and 2 females in the 240 mg/kg group of the satellite group.

Body weight and weight changes:

no effects observed

Food efficiency:

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Inorganic phosphorus and blood potassium concentration were decreased in females in the 240 mg/kg group of the satellite group; these changes were considered to be related to the dysfunction of the digestive tract by the test substance.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

Total excretion volume of potassium and urinary potassium concentration were decreased, urinary pH was lowered and urinary protein was slightly increased in the 240 mg/kg group, total protein, albumin and α1 globulin were decreased and an α2 globulin ratio was increased in males in the 240 mg/kg group of the main study group and inorganic phosphorus was decreased in females in the 60 and 240 mg/kg groups of the main study group; these changes were considered to be related to the dysfunction of the digestive tract by the test substance.

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Histopathologically, blood absorption by the mesenteric lymph nodes was observed in all of the animals which were found dead, and brown pigments in the mesenteric lymph nodes, hemorrhage from the forestomach, edema, polymorphonuclear cell infiltration and inflammation in the glandular stomach in some of them. From these pathological findings, the cause of death was considered to be gastric injury and dysfunction of the digestive tract by the test substance.

The following histopathological findings also confirmed the presence of the gastric injury by the test substance in animals that survived: blood absorption by the mesenteric lymph nodes, brown pigments in the mesenteric lymph nodes and hemorrhage from the forestomach in males or females in the 60 or 240 mg/kg group.

Histopathological findings: neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Squamous cell hyperplasia in the forestomach was noted in 4/4 dead females at 240 mg/kg bw/day.

Squamous cell hyperplasia and an increase in apoptosis in the forestomach, proliferation of surface mucous cell in the glandular stomach in males or females in the 60 or 240 mg/kg group. An increase in mitosis in the glandular stomach was observed in animals of both sexes given 15 mg/kg or more. The latter finding was considered to be an effect of the test substance but was concluded to be toxicologically meaningless because there were no other findings suggestive of gastric injury in those groups.

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

There were no adverse effects on reproductive ability, including delivery and lactation (Tables 17-18, Appedices 19-22).

Dose descriptor:

NOAEL

Effect level:

240 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: No effects

Dose descriptor:

NOEL

Effect level:

240 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: No effects

Clinical signs:

not examined

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Food efficiency:

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

Observation of pups revealed no effects of the test substance on viability, body weights, morphology or necropsy findings (Tables 19-20, Appendices 22-23).

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

240 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: No effects

Dose descriptor:

NOEL

Generation:

F1

Effect level:

240 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: No effects

Reproductive effects observed:

no

Lowest effective dose / conc.:

240 mg/kg bw/day (nominal)

Treatment related:

no

Conclusions:

In a repeated dose and reproduction/developmental toxicity screening test with 1,1'-(1,3-phenylene)bis-1H-pyrrole-2,5-dione in Crl:CD (SD) rats, the NOEL and NOAEL for reproductive/developmental toxicity (parental and F1) were considered to be 240 mg/kg/day.

Executive summary:

In a repeated dose and reproduction/developmental toxicity screening test (C545 (314 -026)), 1,1'-(1,3-phenylene)bis-1H-pyrrole-2,5-dione (99.3%) was administered to four groups of Crl:CD (SD)rats (12 animals/sex/group) by gavage in 0.5% methyl cellulose at dose levels of 0, 15, 60 and 240 mg/kg bw/day 7 days per week for 42 days (males) and 42-52 days (females).

Death occurred in 4 females and 2 males at 240 mg/kg bw/day. These animals showed loose stools, diarrhea, abnormal respiration noise, irregular respiration, etc. before death. In animals that survived, salivation was observed in males in the 60 mg/kg group and in animals of both sexes in the 240 mg/kg group. Total excretion volume of potassium and urinary potassium concentration were decreased, urinary pH was lowered and urinary protein was slightly increased in the 240 mg/kg group, total protein, albumin and α1 globulin were decreased and an α2 globulin ratio was increased in males in the 240 mg/kg group of the main study group and inorganic phosphorus was decreased in females in the 60 and 240 mg/kg groups of the main study group. Inorganic phosphorus and blood potassium concentration were decreased in females in the 240 mg/kg group of the satellite group; these changes were considered to be related to the dysfunction of the digestive tract by the test substance. There were no effects on body weight, food consumption, hematologic parameters, blood coagulation or organ weight.

At necropsy, dilatation of the gastric lumen with gas and/or liquid contents was observed in all animals that died, and brownish mesenteric lymph nodes, red patches in the stomach, dilatation of the lumen of the small intestine with gas and/or liquid red contents and dilatation of the lumen of the large intestine (cecum) with gas contents in some of them. In the pathological examination of animals that survived, brownish mesenteric lymph nodes, black patches in the glandular stomach, dilatation of the intestinal lumen (duodenum, jejunum and cecum) with gas contents and gas contents in the large intestine (colon) were observed, and these were considered to reflect gastric injury and dysfunction of the digestive tract by the test substance.

Histopathologically, blood absorption by the mesenteric lymph nodes was observed in all of the animals which were found dead, and brown pigments in the mesenteric lymph nodes, hemorrhage from the forestomach, edema, polymorphonuclear cell infiltration and inflammation in the glandular stomach in some of them. From these pathological findings, the cause of death was considered to be gastric injury and dysfunction of the digestive tract by the test substance. Squamous cell hyperplasia in the forestomach was noted in 4/4 dead females at 240 mg/kg bw/day. The following histopathological findings also confirmed the presence of the gastric injury by the test substance in animals that survived: blood absorption by the mesenteric lymph nodes, brown pigments in the mesenteric lymph nodes and hemorrhage from the forestomach in males or females in the 60 or 240 mg/kg group. Squamous cell hyperplasia and an increase in apoptosis in the forestomach, proliferation of surface mucous cell in the glandular stomach in males or females in the 60 or 240 mg/kg group. An increase in mitosis in the glandular stomach was observed in animals of both sexes given 15 mg/kg or more. The latter finding was considered to be an effect of the test substance but was concluded to be toxicologically meaningless because there were no other findings suggestive of gastric injury in those groups.

In terms of reproductive/developmental toxicity, there were no adverse effects on reproductive ability, including delivery and lactation. Observation of pups revealed no effects of the test substance on viability, body weights, morphology or necropsy findings.

The NOEL and NOAEL for reproductive/developmental toxicity (parental and F1) were considered to be 240 mg/kg/day, because there were no treatment-related changes in all parameters.

This repeated dose and reproduction/developmental toxicity screening study in the rat is acceptable and satisfies the guideline requirement for this oral study (OECD 422) in rats.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

240 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

The key study was the only study available and was assigned a Klimisch score of 2.

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

The substance N,N′-m-phenylenedimaleimide has been evaluated in one combined repeated dose and reproduction/developmental toxicity screening test in rats

In a combined repeated dose and reproduction/developmental toxicity screening test (OECD 422/GLP), 1,1'-(1,3-phenylene)bis-1H-pyrrole-2,5-dione (99.3%) was administered to four groups of Crl:CD (SD)rats (12 animals/sex/group) by gavage in 0.5% methyl cellulose at dose levels of 0, 15, 60 and 240 mg/kg bw/day 7 days per week for 42 days (males) and 42-52 days (females).

Death occurred in 4 females and 2 males at 240 mg/kg bw/day. These animals showed loose stools, diarrhea, abnormal respiration noise, irregular respiration, etc. before death. In animals that survived, salivation was observed in males in the 60 mg/kg group and in animals of both sexes in the 240 mg/kg group. Total excretion volume of potassium and urinary potassium concentration were decreased, urinary pH was lowered and urinary protein was slightly increased in the 240 mg/kg group, total protein, albumin and α1 globulin were decreased and an α2 globulin ratio was increased in males in the 240 mg/kg group of the main study group and inorganic phosphorus was decreased in females in the 60 and 240 mg/kg groups of the main study group. Inorganic phosphorus and blood potassium concentration were decreased in females in the 240 mg/kg group of the satellite group; these changes were considered to be related to the dysfunction of the digestive tract by the test substance. There were no effects on body weight, food consumption, hematologic parameters, blood coagulation or organ weight.

At necropsy, dilatation of the gastric lumen with gas and/or liquid contents was observed in all animals that died, and brownish mesenteric lymph nodes, red patches in the stomach, dilatation of the lumen of the small intestine with gas and/or liquid red contents and dilatation of the lumen of the large intestine (cecum) with gas contents in some of them. In the pathological examination of animals that survived, brownish mesenteric lymph nodes, black patches in the glandular stomach, dilatation of the intestinal lumen (duodenum, jejunum and cecum) with gas contents and gas contents in the large intestine (colon) were observed, and these were considered to reflect gastric injury and dysfunction of the digestive tract by the test substance.

Histopathologically, blood absorption by the mesenteric lymph nodes was observed in all of the animals which were found dead, and brown pigments in the mesenteric lymph nodes, hemorrhage from the forestomach, edema, polymorphonuclear cell infiltration and inflammation in the glandular stomach in some of them. From these pathological findings, the cause of death was considered to be gastric injury and dysfunction of the digestive tract by the test substance. Squamous cell hyperplasia in the forestomach was noted in 4/4 dead females at 240 mg/kg bw/day. The following histopathological findings also confirmed the presence of the gastric injury by the test substance in animals that survived: blood absorption by the mesenteric lymph nodes, brown pigments in the mesenteric lymph nodes and hemorrhage from the forestomach in males or females in the 60 or 240 mg/kg group. Squamous cell hyperplasia and an increase in apoptosis in the forestomach, proliferation of surface mucous cell in the glandular stomach in males or females in the 60 or 240 mg/kg group. An increase in mitosis in the glandular stomach was observed in animals of both sexes given 15 mg/kg or more. The latter finding was considered to be an effect of the test substance but was concluded to be toxicologically meaningless because there were no other findings suggestive of gastric injury in those groups.

In terms of reproductive/developmental toxicity, there were no adverse effects on reproductive ability, including delivery and lactation. Observation of pups revealed no effects of the test substance on viability, body weights, morphology or necropsy findings.

The NOEL and NOAEL for reproductive/developmental toxicity (parental and F1) were considered to be 240 mg/kg/day, because there were no treatment-related changes in all parameters.

Effects on developmental toxicity

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the available information in the dossier, the substance N,N′-m-phenylenedimaleimide  (CAS No. 3006-93-7) does not need to be classified for reproductive toxicity when the criteria outlined in Annex I of 1227/2008/EC are applied.